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THE  EFFECT  OF  LARGE  APPLICATIONS  OF  COM- 
MERCIAL FERTILIZERS  ON  CARNATIONS 


BY 


FRED  WEAVER  MUNGIE 

A.  B.  Wabash  College,  1910 
M.  S.  University  of  Illinois,  1913 


% 


UNlVER^i^ 


THESIS 

Submitted  in  Partial  Fulfilment  of  the  Requirements 
for  the  Degree  of 

DOCTOR  OF  PHILOSOPHY 
IN  CHEMISTRY 


IN 


THE  GRADUATE  SCHOOL 

OF  THE 

UNIVERSITY  OF  ILLINOIS 


1915 


ACKNOWLEDGMENTS. 
The  author  desires  to  express  his  appreciation  of  the  many  helpful  sug- 
gestions received  from  Dr.  Geo.  D.  Beal,  Dr.  C.  G.   Derick  and  other 
members  of  the  departments  of  Chemistry  and  Botany. 


\ 


THE  EFFECT  OF  LARGE  APPLICATIONS  OF  COM- 
MERCIAL FERTILIZERS  ON  CARNATIONS 


BY 


FRED  WEAVER  MUNCIE 

A.  B.  Wabash  College,  1910 
M.  S.  University  of  Illinois,  1913 


THESIS 

Submitted  in  Partial  Fulfilment  of  the  Requirements 
for  the  Degree  of 

DOCTOR  OF  PHILOSOPHY 
IN  CHEMISTRY 

THE  GRADUATE  SCHOOL 

OF  THE 

UNIVERSITY  OF  ILLINOIS 


1915 


Digitized  by  the  Internet  Archive 

in  2008  with  funding  from 

IVIicrosoft  Corporation 


http://www.archive.org/details/effectoflargeappOOmuncrich 


Effects  of  Large  Applications  of  Commercial  Fer- 
tilizers on  Carnations 

By  Fred  Weaver  Muncie 

In  the  investigation  of  the  use  of  commercial  fertilizers  in  growing 
carnations  by  the  Illinois  Agricultural  Experiment  Station,  it  has  been 
found  that  the  lack  of  appreciation  by  florists  of  the  relatively  high  plant 
food  concentrations  and  often  high  solubilities  of  commercial  fertilizers, 
as  compared  with  manure,  has  often  led  to  a  complete  loss  of  a  crop  of 
flowers  in  an  effort  to  produce  an  extraordinarily  large  one.  On  this 
account,  it  was  considered  desirable  to  study  the  causes  and  effects  of 
overfeeding  with  the  more  ordinarily  used  commercial  fertilizers. 

The  fertilizers  chosen  for  the  experiment  were  dried  blood,  sodium 
nitrate  and  ammonium  sulfate,  acid  phosphate  and  disodium  phosphate, 
and  potassium  sulfate.  For  comparison,  sodium  chloride  and  sodium 
sulfate  also  were  used  on  some  sections.  Experimental  work  upon  the 
subject  was  carried  out  during  the  years  19 12-15. 

Carnations  are  propagated  by  means  of  cuttings,  and  from  these  it  was 
found  impossible  to  secure  a  normal  growth  in  either  sand  or  water  cultures. 
Hence,  the  experimental  work  was  based  upon  the  study  of  plants  grown 
in  soil  carefully  selected  with  the  view  to  securing  uniformity  throughout 
the  benches,  watered  to  give  as  nearly  as  possible  the  same  moisture 
content,  and  subjected  very  nearly  to  identical  conditions  of  heat,  ventila- 
tion, and  illumination.  For  details  regarding  the  type  of  soil,  its  prepara- 
tion, arrangement  of  sections,  etc.,  the  reader  is  referred  to  Bull.  176 
of  the  Illinois  Agricultural  Experiment  Station. 


35707i 


,'  «  ->  J      >  'J    ^     ''' 


The  method  consisted  of  weekly  apphcations  of  the  fertiHzers  at  various 
rates  upon  isolated  sections  in  the  benches,  beginning  about  October  i 
and  continuing  until  about  May  i  or  until  injury  became  serious. 

Effects  of  Overfeeding  on  Condition  of  Plants. — The  rapidity  with 
which  the  sections  of  carnations  became  affected  followed  in  a  general 
way  the  solubility  of  the  fertilizer  used.*  The  solubilities**  of  the  pure 
substances  in  water  per  hundred  parts  at  o°  are  given  in  Table  I. 

Table  I. — Solubilities  op  Pure  vSalts  in  Water  at  o°.     (Parts  per  ioo.) 

NaNO,  (NH4)2S04  NaCl  KCl  K2SO4 

72.9  710  35.7  28.5  8.5 

Na2HP04.i2H20  CaH4(P04)2.H20  CaHP04  CaS04.2H20 

6.3  4  (15°)  0.028  0.241 

Commercial  acid  phosphate  consists  of  about  equal  parts  of  mono-calcium 
phosphate  and  calcium  sulfate.  Reversion  to  monohydrogen  phosphate 
in  presence  of  bases  in  the  soil  would  further  decrease  the  low  solubility 
of  the  acid  phosphate  and  by  double  decomposition  with  calcium,  iron 
and  other  bases  in  the  soil  render  the  sodium  phosphate  first  applied  less 
soluble,  as  pointed  out  by  Cameron  and  Bell.^ 

Dried  blood,  giving  soluble  products  at  a  rate  depending  upon  the 
rapidity  with  which  bacterial  decomposition  proceeds,  could  not  be 
rated  as  having  a  known  solubility  without  a  study  of  the  bacteriological 
activity  of  the  soil  mixture.  Tests  with  litmus  paper  showed  that  the 
surface  of  the  soil,  neutral  at  the  beginning  of  the  experiment,  became 
acid  seven  or  eight  days  after  the  addition  of  the  dried  blood.  Soil  to 
which  ammonium  sulfate  was  applied  became  acid  as  quickly  also. 

Single  applications  of  ammonium  sulfate  and  sodium  chloride  at  the 
rate  of  12.5  kg.  per  100  sq.  ft.  made  on  December  3,  19 13,  produced  marked 
injury  within  a  week's  time.  Equal  amounts  of  potassium  sulfate,  at 
this  time,  followed  by  further  applications  at  intervals  of  one  or  two 
weeks,  at  the  rate  of  1.25  kg.  per  100  sq.  ft.,  produced  no  signs  of  injury 
until  about  January  15,  when  a  lack  of  turgidity  became  noticeable,  fol- 
lowed by  a  gradual  stunting  of  growth,  with  the  more  pronounced  signs 
appearing  only  after  the  middle  of  March.  Signs  of  injury  in  sections 
treated  in  the  same  manner  with  sodium  phosphate  became  evident  even 
more  slowly,  while  acid  phosphate  produced  no  apparent  injury  even  in 
the  largest  applications. 

*  The  impurities  in  the  ammonium  sulfate,  potassium  sulfate  (in  this  case  1.26% 
of  chloride  as  sodium  chloride)  and  disodium  phosphate  are  not  sufficient  to  interfere 
with  the  use  of  the  solubilities  of  the  pure  substances  as  a  rough  measure  of  the  solu- 
bilities of  the  fertilizers  themselves. 

**  Van  Nostrand — Chemical  Annual,  19 10. 


The  fertilizers  may  be  grouped  into  the  class,  easily  soluble  and  pro- 
ducing almost  immediate  injury;  a  second,  moderately  soluble  and  pro- 
ducing delayed  injury;  and  a  third,  difficultly  soluble  and  producing  no 
apparent  injury.  On  days  of  continuous  sunlight  a  more  or  less  pro- 
nounced softness  of  tissue  could  be  detected  by  careful  observation  long 
before  characteristic  injuries  became  apparent. 

Effects  of  Overfeeding  with  Ammonium  Sulfate. — A  marked  softness 
of  tissue  was  the  earliest  sign  of  overfeeding  with  ammonium  sulfate. 
A  complete  plasmolysis  took  place  in  that  portion  of  the  stem  located 
two  and  three  nodes  below  the  bud  and  in  the  portion  of  the  stem  just 
above  the  node,  so  that  the  stem  bent  completely  over.  The  shoots  first 
affected  were  those  with  buds  one-half  to  three-quarters  developed.  At 
the  same  time  white  spots  0.25  and  i.oo  mm.  in  diameter  appeared  upon 
the  upper  leaves  of  these  and  the  younger  shoots.  Microscopic  examina- 
tion of  these  showed  the  chlorophyll  bearing  tissue  entirely  plasmolyzed. 

In  contrast  to  the  injury  from  other  fertilizers,  practically  every  flower 
split.*  This  splitting  was  not  caused  by  the  pressing  outward  of  the 
petals  as  is  usually  the  case,  but  by  a  weakening  of  the  tissue  at  the  line 
joining  the  sepals  to  form  the  calyx  cup.  Later  stages  resulted  in  the 
drying  up  of  the  leaf  tips,  and  the  appearance  of  the  white  depressions 
upon  the  older  leaves.  The  sepal  tips  very  early  became  brown.  Later, 
pustule-like  elevations  about  i  mm.  across  appeared  on  them,  caused 
by  a  crystal  of  ammonium  sulfate  beneath  the  epidermis.  The  injury 
from  excess  of  ammonium  sulfate  was  more  rapid  and  pronounced  in  the 
presence  of  lime  than  without  it. 

Effect  of  Overfeeding  with  Sodium  Nitrate. — Injury  followed  heavy 
applications  of  sodium  nitrate  within  a  few  days,  the  characteristic 
symptom  being  an  even  lightening  of  color  of  the  foliage  over  the  plant, 
followed  by  drying  of  leaf  tips  and  petals  and  withering  of  the  plant. 

Effects  from  Large  Applications  of  Sodium  Chloride.**— The  first  appear- 
ance of  injury  from  large  amounts  of  sodium  chloride  was  two  days  after 
its  application,  a  plasmolysis  of  the  cells  of  the  stem,  causing  it  to  lose  its 
rigidity  at  the  crown.  When  held  within  supports  the  plants  appeared 
normal.  Gradually,  however,  the  plants  lost  their  turgidity  and  the 
chlorophyll  disappeared  evenly  throughout  the  entire  plant.  Tests  made 
in  the  spring  of  19 15  with  heavy  applications  of  sodium  chloride  and 
potassium  chloride  (12  kg.  per  100  sq.  ft.)  showed  the  same  effect  from 
each  of  them,  while  sodium  sulfate,  like  potassium  sulfate,  showed  less 
injury  and  that  only  after  a  longer  period. 

*  Splits  is  a  trade  term  denoting  flowers  with  split  calyces. 
**  Sodium  chloride,  while  not  strictly  a  fertilizer,  was  used  in  the  experiments  be- 
cause of  its  presence  in  considerable  amounts  in  kainite  and  in  some  grades  of  com- 
mercial potassium  sulfate. 


Effects  of  Overfeeding  with  Potassium  Sulfate. — In  earlier  stages 
partial  wilting  occurred  on  days  of  sunshine.  Drying  up  of  the  tips  of  the 
leaves  and  curling  of  the  leaves  upward  upon  their  long  axis  followed, 
with  often,  also,  a  peculiar  inhibition  of  growth  on  one  edge  of  the  leaf, 
with  the  same  on  the  opposite  edge  of  another  portion,  giving  the  leaf  a 
wavy  outline. 

A  marked  stunting  of  growth  was  observable.  This  affected  most 
noticeably  the  lengthening  of  the  stem,  resulting  in  the  later  shoots  assum- 
ing a  rosette  appearance,  due  to  the  leaves  of  normal  length  upon  a  stem 
with  undeveloped  internodes  less  than  an  inch  in  length.  (The  inter- 
node  in  full  grown  shoots  is  ordinarily  three  or  four  inches  long.)  The 
edges  of  the  petals  of  the  flowers  after  about  the  middle  of  January  became 
quite  generally  withered  or  crinkled.  Those  in  the  center  of  the  flower 
remained  closed  quite  tightly,  while  the  other  two  or  three  rows  opened 
normally.  Later,  the  buds  remained  closed,  although  the  pistil  often 
pushed  its  way  out  and  might  be  seen  extending  an  inch  above  the  top 
of  the  bud. 

A  marked  increase  in  exudation  of  nectar  in  the  flower  was  found  to 
have  caused  the  gluing  together  of  the  petals,  and  so  prevented  their 
opening.  On  cloudy  days  very  frequently  a  calyx  cup  would  be  found 
completely  filled  with  this  exudation.  The  exudation  was  most  plentiful 
in  the  flowers  from  plants  receiving  a  moderately  heavy  application  of 
potassium  sulfate  over  a  long  period  of  time  while  the  heavier  applica- 
tions caused  a  noticeable  but  less  plentiful  increase.  A  small  amount  of 
nectar  is  found  in  normal  flowers,  and  somewhat  larger  amounts  in  the 
flowers  from  plants  receiving  large  applications  of  sodium  phosphate, 
sodium  chloride,  ammonium  sulfate,  or  potassium  chloride,  but  not  so 
generally  nor  in  such  large  amounts  as  in  the  sections  treated  with  potas- 
sium sulfate.  Injury  was  less  marked  when  ground  limestone  was  added 
to  the  soil,  in  contrast  to  the  effect  of  liming  on  the  production  of  injury 
by  ammonium  sulfate. 

Effects  of  Overfeeding  with  Sodium  Phosphate. — When  moderately 
large  amounts  of  sodium  phosphate  were  added  over  a  long  period  (as  in 
1913-14)  no  injury  was  noticeable  until  about  the  middle  of  March,  when 
a  retardation  of  growth  was  evident  from  the  decrease  in  height  of  the 
plants  and  abnormally  small  buds  and  flowers.  These  signs  of  inhibition 
became  steadily  more  pronounced  until  the  plants  were  removed  from  the 
benches,  about  May  first.  When  larger  amounts  were  used  (as  12  kg. 
per  100  sq.  ft.  in  1914-15)  loss  of  turgidity  in  the  plants,  longitudinal 
rolling  of  the  leaves,  death  of  the  leaf  tips  and  softness  of  the  petals  of  the 
blossom  were  evident.     These  signs  of  injury  appeared,  however,  only 


after  the  middle  of  January  and  then  only  gradually.     Injury  was  less 
when  the  soil  was  limed  than  when  not. 

Effects  of  Overfeeding  with  Dried  Blood. — In  none  of  the  experiments 
with  dried  blood  did  injury  appear  until  about  the  middle  of  January. 
At  that  time  a  softness  of  the  petals  and  irregularity  of  their  arrangement, 
due  to  the  partial  opening  of  the  inner  and  crinkling  of  the  outer  ones, 
became  more  or  less  common.  The  flowers  became  susceptible  to  brown- 
ing when  a  drop  of  water  from  syringing  lodged  on  a  petal  in  a  position 
to  be  reached  by  the  rays  of  the  sun.  The  height  of  the  plants  was  below 
normal  in  the  spring  but  rather  above  in  the  fall;  the  color  was  good. 
If  the  applications  of  dried  blood  were  not  continued  after  signs  of  injury 
became  apparent,  the  plants  gradually  recovered.  The  same  held  true 
for  plants  overfed  with  ammonium  sulfate  in  contrast  to  those  which  had 
been  injured  by  potassium  sulfate,  sodium  phosphate,  and  sodium  chloride. 

Effects  of  Overfeeding  on  the  Mineral  and  Nitrogen  Content  of  Plants. 
— Effects  upon  the  dry  weight  and  ash  are  shown  in  Table  II,  the  samples 
being  the  foliage  from  the  shoots  gathered  January  9,  1915. 

Table  II. — Dry  Weight  and  Ash  in  Foliage. 


Section 
No. 
269. 

Treatment. 
Check. 

Moist  weight. 
G. 
27.6. 

Dry  weight.    %. 
17.8. 

Ash  (sulfated) 

per  cent,  of  dry 

weight. 

13.68. 

271 

125  P* 

32.4 

17.6 

13-93 

273 

250  P 

32.2 

18.3 

12 

89 

275 

500  P 

30.6 

18.9 

H 

28 

277 

125  K 

26.1 

18.4 

15 

37 

279 

250  K 

36.8 

20.4 

15 

45 

281 

500  K 

32.8 

22.6 

15 

59 

283 

Check 

28.2 

19.2 

13 

19 

285 

125  NaCl 

42.9 

22.8 

14 

45 

The  increase  in  both  values  as  the  applications  of  any  one  fertilizer  in  a 
series  were  increased  is  shown  in  the  table.  The  higher  values  for  plants 
treated  with  potassium  sulfate  and  sodium  chloride  over  those  treated 
with  sodium  phosphate  correspond  to  the  higher  osmotic  pressure  values 
obtained  from  the  sap  of  these  plants  as  well  as  to  the  more  rapid  injury 
from  potassium  sulfate. 

Determination  of  the  total  nitrogen  and  mineral  content  of  the  ash 
from  various  samples  of  plants  treated  with  potassium  sulfate  gave  the 
following  values : 

*  N,  P  and  K  in  the  tables  are  used  to  indicate  ammonium  sulfate,  disodium 
phosphate  and  potassium  sulfate,  respectively,  while  NaCl  indicates  sodium  chloride 
and  A.  P.,  commercial  acid  phosphate.  The  figures  preceding  the  letters  indicate  the 
ntunber  of  grams  applied  weekly  per  20  sq.  ft.  of  bench  space. 


Table  III. — Effect  of  Potassium  Sxh^pate. 

Analyses.     Per  cent. 

Treatment.             NajO.                     KiO.                      SO».  N  (total).  PiO*. 

Check              1.09                 5.38                  1.07  2.58  0.72 

K                     1.25                 6.62                 I. 91  2.53  0.70 


0.16  1.24  0.84  — 0.05  — 0.02 

The  data  show  an  increased  sodium,*  potassium  and  sulfur  content, 
with  practically  a  constant  percentage  of  nitrogen  and  phosphorus. 

A  similar  study  of  plants  to  which  ammonium  sulfate  had  been  ap- 
plied gave  the  results  shown  in  Table  IV. 

Plants  to  which  sodium  phosphate  was  applied  showed  a  higher  phos- 
phorus content,  0.60%  P2O5  and  1.17%  P2O5  in  a  sample  of  1915  in  which 
the  calcium  content  was  decreased  (2.31  and  1.63%  CaO,  respectively, 
in  the  last  set  of  samples) ;  the  nitrogen  content  was  increased  by  applica- 
tions of  sodium  phosphate,  the  values  1.99%,  2.84%  and  3.30%  being 
obtained  from  plants  to  which  had  been  applied,  respectively,  none, 
250  g.  and  500  g.  of  sodium  phosphate  per  20  sq.  ft.  of  bench  space  per 
week  for  several  weeks. 

Table  IV. — Effect  of  Ammonium  Sulfate. 

Analyses.     Per  cent. 


Treatment.  N(total).  N(by  MgO).  SO*.  PiOt. 

Check  2.05  0.168  0.75  0.93 

N  2.93  0.364  2.10  I . 14 


0.88  0.196  1.35  0.21 

The  ratio  2N/SO3  in  ammonium  sulfate  is  28/80  =  0,351,  that  of  total 
nitrogen  to  sulfur  increase  is  0.652;  and  of  nitrogen  by  MgO  0.145.  The 
intake  of  sulfur  when  this  fertilizer  is  used  is  less  than  is  required  for  the 
nitrogen  then,  but  in  excess  of  that  required  to  be  combined  with  the 
nitrogen  determined  by  MgO.**  Limestone  was  found  to  depress  the 
sulfur  intake  from  ammonium  sulfate.  Since  injury  was  greater  in  sec- 
tions so  treated,  the  injury  is  not  proportional  to  the  intake  of  sulfur. 
The  intake  of  phosphorus  was  increased  by  the  addition  of  ammonium 
sulfate,  probably  due  to  acidity  developed  in  the  soil. 

Table  V  shows  the  total  nitrogen  content  of  some  plants  from  Sections 
264  (ammonium  sulfate  and  lime)  and  281  (ammonium  sulfate).     Samples 

*  Mayer^*  states  that  the  addition  of  soluble  potassium  salts  to  a  soil  causes  a 
partial  replacement  of  the  sodium. 

**  The  author  would  not  care  to  report  the  presence  of  ammonium  salts  in  plants 
not  fed  with  it.  It  seems,  rather,  that  MgO  has  caused  some  decomposition  of  the 
organic  material;  the  error  due  to  this  is  assumed  to  be  the  same  in  both  samples. 


were  collected  on  April  25,  1914.  Section  281  had  received  but  one 
application  at  the  rate  of  12.5  kilos  per  100  sq.  ft.  on  December  3,  19 13, 
while  applications  at  the  rate  of  1250  g.  per  100  sq.  ft.  were  made  to  Section 
264  at  15  different  intervals  of  about  two  weeks  after  December  20,  19 13. 
Analyses  were  made  of  upper  and  lower  portions  of  the  plant  separately 
in  order  to  show  any  localization  of  nitrogen  in  the  more  vigorously  grow- 
ing portion  of  the  plant. 

Table  V. — Total  Nitrogen  Determination  on  Foliage. 

%■ 


Sample  No. 

Plant  No. 

Section. 

Portion. 

Condition. 

Nitrogen. 

I 

I 

264-E 

upper 

half  dead 

458 

2 

lower 

half  dead 

4.07 

3 

4 

264-P 

upper 

half  dead 

7.78 

4 

lower 

half  dead 

5.64 

5 

I 

264-P 

upper 

dead 

6.14 

6 

lower 

dead 

3.41 

7 

II 

264-E 

upper 

alive 

6.69 

8 

lower 

alive 

570 

9 

4 

264-E 

upper 

half  dead 

7.01 

10 

lower 

half  dead 

3-34 

II 

11-15 

281-E 

upper 

dead 

4.60 

12 

lower 

dead 

3  02 

13 

.  16 

281-E 

upper 

partially  affected 

4V3 

14 

lower 

partially  affected 

3.78 

15 

20 

281-E 

upper 

half  dead 

4-73 

16 

lower 

half  dead 

2.94 

17 

7 

281-E 

upper 

sUghtly  affected 

4-47 

18 

lower 

slightly  affected 

3.21 

The  total  nitrogen  content  of  the  plants  varied  from  once  and  a  half  to 
more  than  twice  the  normal  value  found  in  the  previous  set.  Average 
values  for  the  plants  from  Section  264  are  6.44%  and  4.43%,  respectively; 
for  those  from  Section  281,  4.63%  and  3.24%.  In  each  case  the  more 
vigorously  growing  portion  contained  the  larger  percentage  of  nitrogen 
and  the  increase  over  the  lower  portion  is  considerably  greater  in  the 
section  to  which  the  smaller  applications  were  made  during  the  entire 
season.  No  clear  relation  is  shown  between  the  nitrogen  content  and 
the  degree  of  injury.  Considerable  tolerance  for  ammonium  sulfate  is 
shown  when  it  was  applied  to  the  soil  in  quantities  not  heavy  enough  to 
produce  immediate,  serious  injury.  The  fact  that  the  dead  plants  had  no 
higher  total  nitrogen  content  than  those  only  injured  is  evidence  that  part 
of  the  nitrogen  when  added  in  small  quantities  was  changed  to  a  nontoxic 
form,  since  the  dead  plants  were  in  this  condition  as  early  as  March  21, 
while  the  living  ones  though  injured  undoubtedly  continued  to  take  up 
the  salt  in  solution  until  samples  were  taken. 

A  series  of  ammonia  determinations  was  made  on  the  sap  from  "checks" 
and  ammonium  sulfate  fed  plants  of  the  set  of  1 2-9-14.  Folin's  micro- 
method  for  the  determination  of  free^*  ammonia  was  used,  the  excess  of 


lO 


sulfuric  acid  (0.01550)  being  titrated  back  with  potassium  hydroxide 
0.02130  with  sodium  ahzarin  sulfonate  as  the  indicator.  Results  are 
given  in  Table  VI. 

Table  VI. — Free  Ammonia  in  Plant  Saps.* 


Sample  No. 

Treatment. 

Appearance. 

Nitrogen. 
Mg.  N  per  cc. 

5 

check 

normal 

none 

8 

250  N 

normal 

0. 1834 

7 

500  N 

normal 

0  1372 

2 

itxxj  N 

slightly  injured 

0.6390 

I 

1000  N 

badly  injured 

I .0560 

The  white  spots  on  the  leaves  of  plants  treated  with  ammonium  sulfate, 
and  of  crystals  imbedded  beneath  the  epidermis  of  the  sepals  were  studied 
by  microchemical  methods.* 

1.  January  21,  19 14.  Plant  Number  4,  Section  281,  White  Enchantress. 
Plant  apparently  normal.  A  drop  of.  sap  from  the  stem  of  a  shoot  was 
treated  with  a  drop  of  ammonia-free  hydrochloric  acid  and  chloroplatinic 
acid,  and  evaporated  at  room  temperature  under  a  loosely  covering  watch- 
glass.  A  few  crystal  masses,  tetrahedral  and  often  aggregated  in  shape 
of  a  cross,  appeared.  They  were  yellow  in  color.  Sap  from  Number  8, 
somewhat  injured,  and  Number  12,  badly  affected,  gave  these  characteristic 
crystals,  also. 

2.  A  section  of  the  leaf  showing  white  blotches  was  immersed  in  chloro- 
platinic acid  after  removal  of  the  epidermis  and  allowed  to  remain  over- 
night. Large  and  perfect  crystals  appeared,  arranged  usually  around 
the  injured  spot,  never  in  it.  They  were  insoluble  in  95%  alcohol  which 
removed  the  excess  of  chloroplatinic  acid. 

3.  A  drop  of  sap  from  plant  Number  4,  Section  281  was  distilled  with  a 
pinch  of  sodium  carbonate  over  a  micro-burner  and  the  distillate  caught 
in  a  hanging  drop  of  hydrochloric  acid  in  a  cover  glass  placed  on  a  glass 
ring  above  it.  Treatment  as  above  gave  small,  yellow  tetrahedra  in- 
soluble in  95%  alcohol. 

Ammonium  salts  were  evidently  present  and  apparently  caused  plas-' 
molysis  of  certain  of  the  chlorophyll  bearing  cells.  Why  injury  of  this 
type  is  caused  by  ammonium  sulfate  in  contrast  to  the  even  lightening 
of  the  color  of  the  whole  leaf  by  the  other  soluble  salts,  sodium  nitrate 
and  sodium  chloride,  is  not  known. 

Nitrate  determinations  according  to  the  phenolsulfonic  method  of 
Mason^^  were  made  upon  the  sap  of  a  "check"  and  an  ammonium  sulfate 
fed  plant  from  the  set  of  March  9,  19 15.  The  values  of  o.oi  and  0.40 
mg.  N  as  nitrate  per  cc.  of  sap,  respectively,  showed  that  nitrification  was 
proceeding  in  the  soil  although  it  was  quite  strongly  acid.^' 

*  In  earlier  stages  of  feeding  with  ammonium  sulfate,  samples  have  been  taken  in 
which  no  NH3  was  detected  by  this  method. 


II 


Total  solids  and  ash  were  determined  on  the  sap  of  the  set  of  1 2-9-14. 
The  results,  given  in  Table  VII,  are  calculated  to  milligrams  per  cc.  of 
sap. 

Table  VII. — Total  Solids  and  Ash  of  Sap. 


Total  solids. 

Ash.* 

Sample  No. 

Set  date. 

Section. 

Treatment. 

Mg.  per  cc. 

Mg.  per  cc. 

2 

12-9-14 

291 

1000  N 

91.9 

3 

293 

1000  K 

104.9 

19.2 

5 

289 

check 

63.8 

II. 8 

6 

261 

check 

62.1 

12  .  I 

7 

265 

250  N 

63.6 

13-9 

8 

267 

500  N 

79-9 

15    I 

9 

277 

125  K 

643 

16. 1 

10 

279 

250  K 

69.9 

17.2 

II 

281 

500  K 

75-7 

17. 1 

12 

283** 

check 

72.1 

150 

I 

I-9-15 

269 

check 

84.0 

7-5 

2 

271 

125  P 

81.7 

132 

3 

273 

250  P 

86.7 

133 

4 

275 

500  P 

93  0 

15    I 

5 

277 

125  K 

92.3 

134 

6 

279 

250  K 

106.3 

20. 1 

7 

281 

500  K 

^    133-7 

20.0 

8 

283** 

check 

105. 1 

14. 1 

The  average  total  solids  content  of  the  sap  was  85 .  i  mg.  per  cc.  and  the 
ash  content  14.9  mg.  The  influence  of  the  fertilizer  applications 
is  seen  in  the  increase  in  both  values  as  the  applications  of  any 
fertilizer  were  increased  in  a  series  of  sections.  Sample  3  of  the  first  set 
and  6  and  7  of  the  second,  all  of  which  were  from  plants  to  which  large 
applications  of  potassium  sulfate  had  been  made,  showed  particularly 
high  values.***  The  first  set  of  data  was  obtained  by  drying  the  samples 
in  a  Sargent  electric  oven  at  60-70°,  the  second  in  a  vacuum  oven  heated 
to  50°  for  12  hours.  The  actual  value  for  total  solids  depended  on  the 
length  of  heating  but  experiments  with  both  sets  of  data  given  showed  the 
same  relative  values  after  several  successive  heatings. 

*  Ash  determinations  upon  the  sap  were  made  by  careful  incineration  of  the  soUds 
in  I  cc.  of  sap  in  platinum  dishes  over  a  low  flame  to  prevent  mechanical  loss  of  particles 
of  the  ash.  The  low  chloride  content  obviates  the  danger  of  volatilization  of  potassium 
chloride  by  high  temperatures. 

**  For  some  reason  total  solids  and  ash  determinations  always  ran  higher  in  sap 
from  plants  in  Section  283  than  from  those  in  other  "check"  sections.  The  same  dis- 
crepancy is  seen  in  the  osmotic  pressure  data  for  these  two  sets. 

***  The  determination  of  total  solids  with  accuracy  is  not  possible  on  account  of  the 
uncrystallizable  solutes  in  the  sap,  and  on  this  account  the  mean  molecular-weight 
calculations  which  often  accompany  osmotic  pressure  data  were  not  made.  Drying 
on  the  water  bath  was  found  to  cause  charring  of  the  sap  from  plants  which  had  been 
treated  with  ammonium  or  potassium  sulfate.  The  first  showed  a  higher  acidity  value, 
the  second  a  higher  sugar  content. 


12 


Determinations  of  sodium  and  potassium  in  the  ash  from  sap  obtained 
on  January  9,  19 15,  from  plants  treated  with  potassium  sulfate,  were  made 
in  order  to  show  the  increased  intake  of  potassium.  Similarly,  determina- 
tions of  phosphorus  were  made  upon  the  sap  from  plants  fertilized  with 
disodium  phosphate.  The  results,  calculated  to  milligrams  per  cc.  of 
sap,  are  given  in  Table  VIII. 

Table  VIII. — Mineral  Content  of  Sap. 


Sample  No. 

Section. 

Treatment. 

NaaO. 
Mg. 

K2O. 
Mg. 

MgrP207 
Mg. 

9 

277 

125  K 

1-4 

9-4 

10 

279 

250  K 

I 

3 

10 

I 

II 

281 

500  K 

I 

3 

10 

I 

12 

283 

check 

I 

2 

8 

4 

I 

269 

check 

I 

5 

2 

271 

125  P 

6 

I 

3 

273 

250  P 

7 

5 

4 

275 

500  P 

9 

6 

Effect  of  Overfeeding  on  Osmotic  Pressure  of  Sap. — Sap  was  expressed 
from  the  stems  of  shoots  after  freezing  them  with  an  ice-salt*  mixture, 
and  the  lowering  of  the  freezing  point  determined  by  the  method  of  Harris 
and  Gortner**  of  allowing  supercooling  until  the  solution  froze  and  cor- 
recting the  value  of  A'  obtained  by  the  formula 

A  =   A'  —  0.0125  mA' 
where  A'  is  the  maximum  temperature  attained  in  the  system  and  u  the 
difference   between    this    value    and    the    minimum    temperature.     The 
relation  between  A  and  the  osmotic  pressure  given  by  Lewis^'  in  the 
approximate  equation 

T    =    I 2 . 06    A 

was  used  in  calculating  the  value  for  t. 

Description  of  Experimental  Method. — Choosing  a  time  when  for  two 
or  more  hours  previous  no  appreciable  draft  had  been  stirring  the  air  in 
the  greenhouse,  from  four  to  eight  shoots  at  the  same  stage  of  growth 
were  removed  from  each  of  the  sections  of  plants  and  quickly  taken  to 

*  Care  was  taken  to  select  samples  from  the  check  and  affected  plants  at  the  same 
time  of  day  and  shoots  in  the  same  stage  of  growth  were  taken,  to  insure  freedom  from 
variations  in  osmotic  pressure  due  to  differences  in  location  and  illumination,  while 
the  fact  that  the  sections  studied  were  usually  adjacent  obviated  the  difficulty  that 
differences  in  temperature  change  the  osmotic  pressure  of  plants.  See  Dixon  and 
Atkins,^'  Atkins,^  Ewart,'^  Drabble  and  Drabble,'^  Cavara.' 

**  The  method  in  general  was  an  adaptation  of  that  recommended  by  Gortner 
and  Harris. ^^"'^  Andre ^  and  also  Dixon  and  Atkins'^  have  shown  that  successive  por- 
tions of  sap  expressed  from  unfrozen  tissue  become  more  concentrated,  while  the  latter 
have  shown  that  the  sap  from  frozen  tissue  always  has  a  lower  freezing  point  than  that 
from  unfrozen,  and  that  successive  portions  gave  nearly  identical  lowerings,  leading 
to  the  conclusion  that  sap  so  expressed  is  representative  of  that  originally  within  the 
tissue. 


13 

the  laboratory.  After  removal  of  the  foliage  from  the  stems,  they  were 
broken  at  the  nodes  and  placed  in  hard  glass  test  tubes  (25  mm.  X  150 
mm.),  stoppered  with  rubber  stoppers  and  sealed  with  oil  paper  and  rub- 
ber bands.  Freezing  was  produced  by  the  use  of  the  ice  and  salt  bath,* 
giving  a  temperature  of  — 15°  or  lower  and  allowing  the  tubes  to  remain 
in  the  refrigerator  overnight.  The  tubes  were  then  removed  from  the  bath 
and  after  the  walls  had  been  cleaned  with  distilled  water  and  wiped  dry, 
the  portions  of  shoots  were  removed,  thawed  gradually,  and  the  sap  ex- 
pressed by  pressure  from  the  screw  of  a  tincture  press  set  perpendicular 
to  the  wall  upon  two  pieces  of  Vs  inch  plate  glass.  After  a  first  expression, 
the  shoots  were  rearranged  and  pressure  again  applied.  The  sap  was 
filtered  through  an  S.  &  S.  589  filter — with  a  watch  glass  over  the  funnel 
to  minimize  evaporation — into  a  small  test  tube;  a  drop  of  xylene  was 
added  as  a  preservative  and  the  tubes  placed  at  once  in  a  refrigerator,  at 
about  10°.  The  sap  after  filtration  was  usually  a  clear,  brown  liquid 
without  sediment. 

As  soon  as  convenient  the  freezing-point  determinations  were  made. 
A  thermometer  was  used  having  a  bulb  about  5  mm,  by  35  mm.,  the  mer- 
cury tube  enclosed  in  a  hollow  jacket,  and  graduated  to  — 6 . 5  °  in  tenths 
of  degrees,  upon  which,  by  the  aid  of  a  lens,  hundredths  of  a  degree  could 
be  read  without  danger  from  parallax.  A  stirrer  of  platinum  wire  and 
the  thermometer  were  placed  in  the  5  cc.  of  sap  contained  in  a  test  tube 
of  Bohemian  glass  (15  X  120  mm.)  and  the  whole  cooled  to  about  +2° 
in  an  ice  and  salt  bath  in  a  beaker.  The  tube  was  wiped  free  from  water 
and  placed  within  a  hard  glass  test  tube  (25  mm.  X  150  mm.)  set  two- 
thirds  way  into  the  ice  and  salt-freezing  mixture.  It  was  found  saving 
of  time  to  place  this  bath  in  a  Dewar  bulb,  with  inside  diameter  of  35  X 
130  mm.;  the  top  was  closed  with  a  piece  of  cork;  the  bath  so  arranged 
remaining  effective  for  three  hours  or  more  of  use.  During  the  entire 
cooling,  the  sap  was  constantly  stirred  to  prevent  its  freezing  about  the 
sides  of  the  tube.  The  lowest  temperature  obtained  was  read  to  one- 
tenth,  and  the  maximum,  by  the  aid  of  a  lens,  to  one-hundredth  degree. 
The  tube  was  removed  to  a  beaker  of  water,  and  after  the  temperature 
had  risen  to  about  10°,  the  determination  duplicated  to  within  0.01°, 
usually  without  difficulty  on  the  first  trial.  A  typical  determination 
gave  the  following  values: 

A'  =  1.28  u  =  4.12  A  =  1. 214 

A'  =   1.27  «  =  3.43  A  =   1. 216 

Average  1.2 15  from  which  tt  =  14.64  atmospheres. 

*  It  was  found  cShvenient  in  case  less  than  a  dozen  tubes  of  material  were  frozen* 

to  place  the  ice  and  salt  bath  in  one  or  two  one-liter  Jena  beakers.     In  this  way  the 

ice  can  be  packed  about  the  upper  portions  of  the  test  tubes,  and  the  beakers,  with  fire 

or  six  test  tubes  in  them,  are  narrow  enough  to  keep  the  tops  of  the  test  tubes  from 

touching  the  solution. 


14 


Table  IX. — Osmotic  Pressure  Determinations. 

Sample 

Section.      Treatment.  A'. 

3-21 
403 

3  90 
1 .  10 

567 
3  40 
4.80 
5  80 
2.87 
4-43 

5-34 
4.78 
5  30 
4-55 

3-91 
5  10 
5  40 

3-75 
4.82 
432 
4.14 

4.00 
4.62 
4.98 
3.81 
2.65 
4.92 

463 
4.92 

2.92    1. 77 1    21.40 
5.52    1.338    16.13 

)Sin  be  made  between  the 
values  for  the  osmotic  pressure  determined  in  successive  sets  on  account 
of  variations  due  to  temperature,  physiological  scarcity  of  water,  etc., 
but  the  values  obtained  from  plants  in  adjacent  sections  at  the  one  time 
are  regular  enough  to  be  comparable. 

From  the  values  for  osmotic  pressure  of  Samples  7,  8,  2  and  i  of  the  set 
of  1 2-9-14  the  conclusion  was  drawn  that  the  osmotic  pressure  within 
the  plants  increased  as  the  quantity  of  ammonium  sulfate  applied  to  the 
soil  was  increased.  Samples  2,  3  and  4,  and  5,  6  and  7  of  the  set  of  1-9- 15 
gave  similar  results  with  increasing  applications  of  sodium  phosphate 
and  potassium  sulfate.  The  values  obtained  from  the  application  of 
sodium  phosphate  were  in  every  case  lower  than  those  obtained  from  appli- 
cation of  equal  quantities  of  potassium  sulfate  or  ammonium  sulfate.     The 


Date. 

No. 

Section. 

Treatment. 

A'. 

II-12-15 

I 

291 

1000  N 

1.30 

- 

2 

293 

1000  K 

1-37 

3 

295 

1000  P 

1.32 

4 

289 

check 

1 .15 

5 

269 

check 

1 .00 

II-20-14 

I 

291 

1000  N 

1-33 

2 

293 

1000  K 

1.50 

3 

295 

1000  P 

1 .  10 

4 

289 

check 

1 .20 

5 

283 

check 

1. 18 

7 

283 

check 

1.27 

12-9-14 

I 

291 

1000  N 

1.66 

(10  A.M.) 

2 

291 

1000  N 

1-43 

3 

293 

1000  K 

1 .40 

5 

289 

check 

0.95 

6 

261 

check 

0.99 

7 

265 

250  N 

1 .  10 

8 

267 

500  N 

1.28 

I2-9-I4 

9 

277 

125  K 

1.05 

(4  P-M.) 

10 

279 

250  K 

1. 18 

II 

281 

500  K 

1. 18 

12 

283 

check 

1 .06 

I-9-15 

I 

269 

check 

1 .20 

2 

271 

125  P 

1.28 

3 

273 

250  P 

1.32 

4 

275 

500  P 

1-39 

5 

277 

125  K 

I   35 

6 

279 

250  K 

1.58 

7 

281 

500  K 

1.87 

8 

283 

check 

1.28 

9 

285 

125  NaCl 

1.88 

10 

287 

500  A.  P. 

1.48 

Discussion 

of  Results.- 

-No   comp 

arisoE 

A. 

T. 

I  .210 

14.60 

I  .261 

1521 

I    195 

14.41 

I    054 

12.71 

0.946 

II  .41 

I  .196 

14.42 

1.396 

16.84 

0.994 

11.99 

1.078 

1300 

I  .098 

13     24 

I  .160 

13   99 

1.513 

18.24 

I    305 

15.73 

I  .267 

15.25 

0.856 

10.34 

0.901 

10.86 

0.990 

11.94 

I.  174 

14. 16 

0.962 

II  .60 

0.973 

11.73 

I  .076 

1301 

0.967 

11.68 

I  .100 

13.24 

I  .  169 

14.08 

I.  178 

14.20 

1.284 

15.50 

1.265 

15.29 

1.448 

17.49 

I  .722 

20.76 

I  .161 

14.04 

15 

samples  taken  on  11-12-14  and  11-20-14  gave  higher  values  for  the  sap 
from  plants  overfed  with  potassium  sulfate  than  those  treated  with  am- 
monium sulfate,  but  later  in  the  year  in  the  set  of  12-9- 14  (Samples  i 
and  2)  the  relative  values  are  reversed. 

In  the  set  of  1 2-9-14  plants  treated  with  potassium  sulfate  at  the  rate 
of  1000  g.  per  section  per  application  were  still  apparently  normal,  al- 
though the  osmotic  pressure  amounted  to  15.25  atmospheres,  while  plants 
treated  with  one-half  this  weight  of  ammonium  sulfate  possessed  an  os- 
motic pressure  of  only  14.16  atmospheres  and  showed  signs  of  injury. 
Injury,  on  the  other  hand,  had  not  appeared  on  plants  treated  with  am- 
monium sulfate  (250  g.  per  section  per  application)  when  the  osmotic 
pressure  amounted  to  12.42  atmospheres  as  compared  to  11.34  atmos- 
pheres in  the  adjacent  "check"  section  (12-9-14 — 10  a.m.). 

The  higher  value  of  Sample  i  over  Sample  2  (of  the  set  of  1 2-9-14 — 
10  A.M.)  was  correlated  with  a  greater  degree  of  injury  by  the  ammonium 
sulfate.  Injury  appeared  on  the  plants  from  sections  to  which  potassium 
sulfate  was  applied,  only  when  an  osmotic  pressure  of  over  twenty  atmos- 
pheres was  reached  (1-9-15),  and  an  osmotic  pressure  value  up  to  15. 50 
atmospheres  was  found  in  plants  on  soil  treated  with  sodium  phosphate, 
without  injury  being  apparent.  The  determination  of  the  value  on  the 
sap  from  plants  treated  with  acid  phosphate  gave  1 6 . 1 1  atmospheres, 
yet  these  plants  exceeded  in  size  and  vigor  those  to  which  no  fertilizer 
was  applied  ('1-9-15).  The  conclusion  to  be  drawn  from  these  facts 
is  that,  with  a  single  fertilizer,  injury  from  overfeeding  becomes  apparent 
when  a  certain  osmotic  pressure  is  reached,  but  that  this  value  is  different 
for  different  fertilizers. 

The  injury  from  applications  of  sodium  chloride  at  the  rate  of  125  g. 
per  section  per  application,  occurred  at  approximately  the  same  time, 
was  very  similar  to,  and  was  of  about  the  same  degree  as  that  from  ap- 
plications of  potas.sium  sulfate,  in  four  times  these  quantities.  The  rela- 
tive osmotic  pres.sure  values  are  given  in  Samples  9  and  10  (1-9-15). 
The  solubilities  of  these  salts,  as  pointed  out  on  page  2785,  at  0°  are 
35.7  and  8.5,  respectively,  giving  a  ratio  roughly  of  4  to  i. 

Effects  of  Overfeeding  on  the  Total  Acidity  of  the  Cell  Sap. — Reaction 
tests  with  litmus  paper  showed  that  the  soil  receiving  no  fertilizer  or 
only  manure  was  neutral  or  slightly  alkaline  in  the  fall,  and  that  a  gradual 
change  to  slight  acidity  took  place  during  the  winter.  Commercial 
acid  phosphate,  dried  blood  and  ammonium  sulfate  upon  the  soil  each 
increased  the  total  acidity,*  the  first  one  immediately  after  application, 

*  It  is  not  likely  that  the  hydrogen-ion  concentration  of  the  soil  solution  was  greatly 
increased  by  addition  of  commercial  acid  phosphate,  since  the  formula  used  in  its  prep- 
aration prevents  the  presence  of  free  sulfuric  acid  by  providing  a  slight  excess  of  tri- 
calcium  phosphate.  Salm.'^^  using  a  hydrogen  electrode  apparatus,  found  [H]  =  3.3  X 
io~'  for  the  di-hydrogen  sodium  phosphate  at  18°  in  o.i  N  sol. 


i6 


the  latter  two  within  about  a  week's  time.  In  the  case  of  these  fertilizers, 
the  surface  of  the  soil  became  acid  after  the  lower  portions.  When  di- 
sodium  phosphate  was  applied,  the  surface  of  the  soil  became  alkaline 
to  litmus,  the  deeper  parts  becoming  alkaline  more  slowly.  Tests  on 
Section  275  (500  P)  on  February  18,  1915,  and  on  291  (1000  P)  on  March 
22,  19 15,  showed  that  the  soil  at  each  successive  inch  to  the  bottom  of  the 
bench,  was  alkaline  to  litmus.  In  so  far  as  could  be  determined  by  this 
method,  applications  of  potassium  sulfate  and  of  sodium  chloride  did  not 
change  the  reaction  of  the  soil.*  Hence  an  opportunity  was  given  to  study 
the  effect,  upon  the  acidity  of  the  cell  sap,  of  fertilizers  producing  increased 
acidity  in  the  soil,  alkalinity,  and  no  change  in  reaction,  and  upon  the  re- 
lation the  changes  bore  to  injury  from  overfeeding  with  the  fertilizer. 

Determinations  were  made  by  titrating  at  about  15"  with  C02-free 
KOH,  approximately  0.02  N,  i  cc.  portions  of  sap  diluted  to  6  cc.  with 
C02-free  water,  using  phenolphthalein  as  the  indicator.  Results  are  cal- 
culated as  cc.  of  normal  acid  per  cc.  sap.** 

Table  X. — Acidity  op  Plant  Sap.*** 


Sample 

Condition 

Date. 

No. 

Section. 

Treatment. 

of  plants. 

Cc.  N  acid. 

12-10-14 

I 

291 

1000  N 

affected 

0 .  03068 

2 

293 

1000  K 

normal 

0.02492 

3 

295 

ICXX)  P 

normal 

0.05490 

4 

289 

check 

normal 

0 . 02048 

5 

261 

check 

normal 

0 . 02090 

6 

265 

250  N 

normal 

0.02238 

7 

267 

500  N 

affected 

0.02728 

8 

277 

125  K 

normal 

0.02088 

9 

279 

250  K 

normal 

0.02002 

10 

281 

500  K 

normal 

0.02130 

II 

283 

check 

normal 

0.02130 

I-14-15 

12 

269 

check 

normal 

0.01977 

13 

271 

125  P 

normal 

0.05035 

14 

273 

250  P 

normal 

0 . 06438 

15 

275 

500  P 

normal 

0.07415 

16 

277 

125  K 

normal 

0.02319 

17 

279 

250  K 

normal 

0 . 02039 

18 

281 

500  K 

affected 

0.02422 

19 

283 

check 

normal 

0.02252 

20 

285 

125  NaCl 

affected 

0.01870 

21 

287 

500  A.  P. 

vigorous 

0.06981 

*  See,  however,  Maschaupt.'^" 
**  For  memoir  on  acidity  in  plants,  see  Astruc* 
***  Boiling  a  solution  of  CO2  in  distilled  water  under  diminished  pressure  by  warm- 
ing the  test  tube  with  the  hand  was  found  completely  to  remove  the  CO2.     Similar  treat- 
ment of  sap  gave  identical  values  for  acidity  before  and  after.     Hence,  the  acidity 
was  not  due  to  dissolved  CO2. 


17 

Acidity  values  remained  about  the  same  when  potassium  sulfate  was 
appHed,  but  increased  after  appHcations  of  acid  phosphate,  ammonium 
sulfate  or  disodium  phosphate,  being  proportional  in  each  case  to  the 
amount  put  on  the  soil.  The  increased  total  acidity  following  applications  of 
disodium  phosphate  (which  is  alkaline  to  phenolphthalein)  was  unexpected 
and  a  more  detailed  study  was  made  of  the  sap  from  these  plants.  Ether- 
soluble  acids  were  absent  and  none  of  the  phosphate  was  extracted  by 
moisture-free  ether.  Phosphate  was  determined  in  i  cc.  portions  of 
Samples  12-15  and  the  total  acidity  of  the  solution  calculated  on  the  as- 
sumption of  the  phosphorus  being  present  (i)  as  orthophosphoric  acid, 
and  (2)  mono-alkali  phosphate,*  the  values  being  given  in  Table  XI. 
Table  XI. — Acidity  of  Sap  by  Titration  and  Calculation. 

Acidity. 


Sample 
No. 

Treatment. 

Mg2P207. 

As  H3PO4. 

As  XH2PO4. 

By  titration. 

12 

check 

0.0015 

0.02692 

0.01346 

0.01977 

13 

125  P 

0.0061 

0. 10768 

0.05384 

0.05035 

/    14 

250  P 

0.0075 

0. 13460 

0.06730 

0.06438 

15 

500  P 

0.0096 

0.16348 

0.08174 

0.07415 

The  values  calculated  as  XH2PO4  agree  more  closely  than  those  for 
H3PO4,  pointing  to  the  presence  of  the  phosphate  as  mono-alkali  phos- 
phates. Subtraction  of  the  "check"  value  for  Mg2P207  from  each 
of  the  other  values  to  obtain  the  increase  in  phosphate  intake  due  to 
applications  of  disodium  phosphate  and  comparison  of  the  titratable 
acidity  calculated  from  these  results  with  the  excess  of  acidity  of  the  solu- 
tions over  that  of  the  "check"  gives  the  following  results: 

Table  XII. — Acidity  and  Phosphorus  Content  Due  to  Overfeeding. 

Increase  in  P2O5. 


(1)     As  Mg2P207. 

(2)    As  MgmH. 

Titration. 
(3)   As  MgmH. 

Ratio. 

(3)/ (2). 

0 . 0046 

0.03999 

0 . 03058 

0.765 

0 . 0060 

0.05388 

0.04461 

0.827 

0.0081 

0.07276 

0.05438 

0.747 

The  ratio  between  the  value  of  H  determined  by  titration  and  by  the 
gravimetric  method  at  15°  was  determined  to  be  0.905,  so  that  the  ratios 
obtained  are  in  the  same  direction,  although  the  lower  values  for  the  sap 
indicate  that  some  of  the  phosphate  may  have  been  present  as  the  mono- 
hydrogen  phosphate. 

This  method  was  applied  to  the  problem  of  determining  the  salt  in  form  of  which 
phosphorus  enters  the  plants.  In  every  case  increasing  applications  of  disodium  phos- 
phate gave  higher  acidity  values.  When  brown  rock  phosphate  was  used  (nasturtiums 
grown  in  sand  culture  with  Hopkin's  nutrient  solution  omitting  phosphorus  after  the 
first  application)  a  regular  increase  up  to  a  maximum  in  size  of  plants  followed  by  a 

*  Two  hydrogens  of  orthophosphoric  acid  and  one  of  monosodium  phosphate 
when  the  solution  is  concentrated  at  0°  and  phenolphthalein  is  the  indicator."'  At 
higher  temperatures,  hydrolysis  of  the  salt  increases  the  alkalinity  of  the  solution. 


i8 

decrease  was  obtained,  without  a  consistent  variation  in  the  acidity  of  the  sap.     Rock 
phosphate  apparently  is  not  taken  into  the  plant  as  mono-calcium  phosphate. 

Reaction  of  the  soil  to  litmus  paper  was  determined  from  time  to  time. 
After  the  first  applications  of  sodium  phosphate  the  soil  reacted  alkaline 
to  litmus  on  the  surface,  with  decreasing  alkalinity  or  acidity  as  the.  dis- 
tance below  the  surface  increased.  On  March  22,  191 5,  Section  295 
(to  which  applications  of  1000  g.  of  sodium  phosphate  had  been  made) 
was  found  to  have  an  alkaline  reaction  to  litmus  paper  when  tested  for 
each  inch  of  soil  down  to  the  bottom  of  the  bench  (5  inches).  Two 
shoots  each  from  plant  Number  4,  badly  injured,  and  plant  Number  12, 
apparently  normal,  were  taken  and  the  sap  expressed  without  previous 
freezing.     The  sap  reacted  acid  to  phenolphthalein  in  each  case. 

The  power  of  soils  to  absorb  bases  from  salts  is  well  known. ^  With 
this  in  mind,  a  liter  of  solution  of  disodium  phosphate  was  made  up  with 
carbon  dioxide-free  water,  and  aliquot  portions  titrated  with  standard 
sulfuric  acid  to  a  faint  rose  coloration,  using  phenolphthalein  as  the  indi- 
cator. Six  carnation  cuttings,  rooted  in  water,  were  cleansed  by  repeated 
washing  with  distilled  water  and  floated  on  the  surface  of  500  cc.  of  the 
solution  by  placing  them  in  holes  of  a  paraffined  cork.  They  were  placed 
in  the  greenhouse  for  six  days,  covered  with  a  large  bell  jar  and  shaded 
during  the  daytime.  The  cuttings  were  taken  out,  the  solution  care- 
fully rinsed  off  and  after  removal  of  the  roots  the  remainder  of  the  shoots 
was  frozen,  the  sap  expressed,  and  i  cc.  portions  titrated  with  standard 
alkali,  using  phenolphthalein  as  the  indicator.  Comparison  was  made 
with  the  acidity  of  the  sap  from  cuttings  taken  from  the  cutting  bench 
and  prepared  as  in  the  former  case  for  sap  expression. 

Strength  of  Solution  2  G.  Na2HP04.i2H20  per  Liter. 

Titration  of  10  cc.  portions  Titration  of  plant  sap. 

HsS04  (0.01550  AT).  KOH  (0.02130  N). 


(1).  (2).  (1)  Check.  (2)  Treated. 

0.32  cc.  0.31  CC.  0.92  cc.  1. 3 1  CC. 

0.0048  CC.  A'  alkali  per  cc. 

In  the  absence  of  soil,  the  sap  had  become  more  acid  when  the  plants 
were  grown  in  the  disodium  phosphate  solution,  hence  the  increased 
acidity  could  not  be  attributed,  at  least  entirely,  to  the  absorptive  power 
of  the  soil  for  bases. 

Effect  of  Large  Applications  of  Potassium  Sulfate  on  Carbohydrate 
Content  of  Sap  and  Foliage. — The  increased  exudation  of  nectar  and 
gluing  together  of  the  petals  in  the  flowers  on  plants  which  had  been  treated 
with  large  amounts  of  potassium  sulfate  has  been  listed  among  the  charac- 
teristic signs  of  overfeeding  with  this  fertilizer  (page  6).  An  attempt 
was  made  to  determine  the  cause  of  this  increased  flow. 

The  amount  of  nectar  present  in  an  aff'ected  flower  amounted  to  as  much 
as  I  cc.  in  the  spring  of  191 2-13,  when  applications  of  potassium  sulfate. 


19 

moderate  when  compared  with  those  used  in  19 14-15,  were  made  weekly 
during  the  season  October  to  May.  In  19 14-15  the  flow  was  not  so  plenti- 
ful, although  noticeably  greater  than  in  the  "check"  flowers.  In  the  for- 
mer year,  the  nectar  was  a  brownish  liquid  with  a  sweet  and  bitter  taste, 
miscible  with  water,  while  in  the  latter  year  it  was  a  clear,  colorless 
liquid.  It  had  a  sweet  taste,  and  was  neutral  to  litmus  and  phenolphthal- 
ein.  It  charred  on  ignition  on  a  platinum  foil,  with  the  odor  of  burnt 
sugar,  leaving  a  small  amount  of  ash  which  was  alkaline  to  moist  litmus 
paper  and  to  phenolphthalein.  Sodium  and  potassium  flame  tests  were 
positive,  calcium  doubtful.  No  indication  of  tannin  was  given  by  tests  with 
neutral  ferric  chloride  and  with  potassium  ferricyanide  and  ammonia. 
A  solution  made  by  washing  off  the  nectar  with  distilled  water  reduced 
Fehling's  solution.  A  heavy  osazone  precipitate  of  bright  yellow  color 
was  thrown  down  upon  heating  it  in  a  boiling  water  bath  with  phenyl- 
hydrazine,  acetic  acid  and  a  crystal  of  sodium  acetate,  after  three  minutes' 
boiling.  Ten  minutes'  boiling  increased  the  amount.  A  much  heavier 
osazone  precipitate  was  given  after  a  few  minutes'  boiling  with  hydro- 
chloric acid,  and  a  portion  of  the  solution  inverted  by  the  Clerget  method 
gave  a  heavier  osazone  precipitate  than  a  similar  amount  before  inver- 
sion. The  rotation  in  a  i  dm.  tube  of  1.5°  Ventzke  was  changed  to  1.18° 
V.  after  the  Clerget  inversion.  Hence,  glucose  and  sucrose  were  present. 
The  precipitate  formed  in  the  hot  solution  was  filtered  off  and  the  filtrate 
again  boiled  till  no  further  precipitate  separated.  On  cooling  the  fil- 
trate a  further  precipitate  of  sodium  acetate  and  osazone  separated. 
This  osazone  possessed  a  roset  structure  characteristic  of  maltosazone, 
and  was  soluble  in  the  boiling  solution  and  reprecipitated  from  it  on 
cooling  as  is  maltosazone.  Not  enough  of  the  precipitate  could  be  ob- 
tained after  recrystallization  for  a  melting-point  determination.*  Tests** 
made  with  a  guaiacol  solution  and  neutral  hydrogen  peroxide  gave  a  nega- 
tive test  with  the  exudation,  but  an  equally  intensive  color  with  sections 
of  petal,  ovary,  leaf  and  stem  of  both  normal  and  affected  plants.  Neither 
of  the  reagents  used  alone  gave  a  reaction.  Microscopic  examination  of 
the  lower,  plasmolyzed  portions  of  the  petals  showed  the  cell  walls  intact 
and  of  normal  thickness.  It  was  concluded  from  this  that  the  increased 
amount  of  sugar  was  not  due  to  breaking  down  of  these  cell  walls,  but  was 
an  exudation.  Experiments  were  then  undertaken  to  compare  the  sugar 
content  of  the  sap  expressed  from  the  stems  of  the  plants  not  fertilized  and 
of  those  receiving  applications  of  potassium  sulfate.  Evidence  that  a 
larger  amount  of  sugars  was  present  in  the  sap  of  the  latter  plants  was 

*  Brown  and  Morris*  used  200  g.  of  leaf  tissue  in  order  to  obtain  enough  for  prep- 
aration of  maltosazone. 

**  Griiss'^  believed  gummosis  might  be  caused  by  an  excess  of  diastatic  enzyme 
and  used  this  reagent  as  a  means  of  detecting  it. 


20 


found  during  the  determination  of  total  solids  of  the  sap  {vide  supra), 
when  the  residue  from  this  sap  was  of  greater  weight  and  charred  at  a 
lower  temperature  than  that  of  the  check. 

The  comparative  optical  rotations*  and  copper-reducing  powers  of 
sap  from  "check"  sections  and  those  which  had  received  applications  of 
potassium  sulfate  are  shown  in  Table  XV. 

Table  XV. — Optical  Rotation  and  Cu-Reducing  Power  of  Sap  Solutions. 


Rotation 
circ.  degrees. 


Reducing  power. 
Mft.     CuO. 


Date. 
1-9-15° 


2-IO-15 


2-17-I5 


3-9-15 


Treatment. 

check 
125  K 
250  K 
500  K 
check 
K 
check 
250-500  K 
check 
250-500  K 


Orig. 

0.73 
1. 91 
1.42 
1.49 
3-23 
3-51 
2.81 
3.26 

2-43 
3.00 


Hydrolyzed.*    Complete.*    Orig. 
0.83 

1-25 

0.97 

I  .21 


Clerget.     Comp 


I  .20 

1-35 

1-53 
2.28 

1-43 
1. 91 


67 
79 
06 
34 


556 
476.5 
521 
522 


1434 
1461 

1276 

1273 


1654 
1976 

(1282) 

1438 


In  view  of  the  work  of  Davis,  Daish  and  Sawyer,^  it  seems  possible, 
though  not  proven,  that  the  quantitative  relationships  of  the  sugars 
in  expressed  sap  may  not  represent  the  condition  within  the  living  tissue. 
The  consistently  higher  values  obtained  by  both  methods  of  estimation, 
showed,  however,  that  the  application  of  potash  to  the  soil  had  resulted 
in  an  increased  carbohydrate  production,  in  a  more  rapid  hydrolysis  of 
starch,  or  in  a  greater  permeability  of  the  cell  membranes  in  the  meso- 
phyll  tissue,  so  that  a  larger  amount  of  sugar  was  found  within  the  conduc- 
ing and  storage  tissues. 

Leaf  tissue  (Set  2-1 0-15)  dried  at  50-70°  was  extracted  with  80% 
alcohol  (i  g.  pptd.  CaCOs  being  added  to  neutralize  acids  present)  and 
the  extracts,  after  removal  of  alcohol,  cleared  with  5  cc.  neutral  lead  ace- 
tate, I  cc.  basic  lead  acetate  and  alumina  cream.     The  extracts  from  7  g 

*  A.  Schmidt  and  Hausch  half -shadow  polariscope,  with  tubes  4  dm.  long,  was 
used.  CuO  values  were  obtained  by  using  Defren's^"  solution,  the  copper  being  de- 
termined by  Low's  method  (Treadwell  and  Hall,  p.  682). 

°  5  cc.  sap  diluted  to  50  cc.  cleared  with  5  cc.  basic  lead  acetate  (sp.  gr.  1.115) 
and  an  excess  of  alumina  cream. 

*  20  cc.  sap  diluted  to  100  cc.  cleared  with  10  cc.  basic  lead  acetate  and  alumina 
cream. 

'  10  cc.  sap  diluted  to  100  cc.  with  5  cc.  basic  lead  acetate  and  alumina  cream. 
"^  10  cc.  sap  diluted  to  100  cc.  with  2  cc.  basic  lead  acetate  and  alumina  cream. 

*  Hydrolyzed  24  hours  with  10%  0.5  N  HCl  at  70°. 
^  Clerget  inversion. 

"  Inversion  for  3  hours  in  boiling  water  bath  of  25  cc.  soln.  12V2  cc.  water  and 
2.5  cc.  HCl  sp.  gr.  1. 19. 


21 

made  up  to  loo  cc.  gave  values  shown  in  Table  XVI.     A  trace  only  of 
pentoses  was  found  in  the  extract. 

Table  XVI. — Sugar  Determinations  in  Extracts. 

Cupric-reducing  power. 
Mg.  CuO. 


Section. 

Treatment. 

268-270 

check 

277 

125  K 

Original.  Clerget.  Complete. 

398.0  1656.8  1933-6 

652.8  1873.6  1990.4 

The  results  are  similar  to  those  in  Table  XV. 

Examination  was  made  for  starch  in  carnation  leaves  taken  from  the 
plant  after  a  day  of  sunshine  by  boiling  them  for  some  time  in  alcohol, 
then  in  water,  and  testing  leaf  sections  with  an  alcoholic  solution  of  iodine ; 
starch  was  found  to  be  plentiful.  Comparative  determinations  of  the 
starch  content*  were  made  upon  the  residues  from  sugar  extractions,  using 
a  diastase  solution  prepared  by  extraction  of  ground  malt  with  mono- 
sodium  phosphate  solution  at  ice-box  temperature,  but  not  dialyzed.** 
Fifty  cubic  centimeters  of  water  were  added  to  the  residue  and  the  starch 
gelatinized  by  boiling  for  five  minutes,  with  continuous  stirring.  After 
cooling  to  60°,  5  cc.  of  the  diastase  solution  were  added  with  a  pipet  and 
digestion  allowed  to  proceed  for  an  hour.  The  mixture  was  again  heated 
to  boiling  and  5  cc.  of  diastase  again  added  and  after  an  hour  the  mixture 
was  filtered  and  washed  thoroughly.  The  maltose  in  the  filtrate  was 
hydrolyzed  to  glucose  by  the  modified  Sachsse  method  and  glucose  deter- 
mined with  Fehling's  solution,  correction  being  made  for  maltose  in  the 
diastase  solution.  The  values  obtained  for  samples  from  sets  of  2-10-15 
and  2-1 7-1 5  are  shown  in  Table  XVII. 

Table  XVII. — Starch  Content  of  Carnation  Leaves. 

Starch  per  cent. 
Treatment.  2-10-15.  2-17-15. 

check  2.72  3-44 

K  1.94  3.09 

A  lower  starch  content  in  "check"  tissue  is  indicated  by  the  results. 
While  these  analyses  were  not  made  over  a  long  enough  period  to  form 
a  basis  for  a  conception  of  the  efi"ect  produced  by  potash  upon  carbohydrate 
production  and  transformations,  the  higher  sugar  with  lower  starch 
content  is  interesting  in  view  of  the  work  of  Sherman  and  Thomas^^  upon 
the  activating  action  of  potassium  sulfate  upon  diastase. 

Summary. 

The  purpose  of  the  investigation  was  to  determine  the  effects  upon  the 
plants  of  large  applications  of  certain  commercial  fertilizers  to  the  soil 
on  which  carnations  were  grown. 

*  Brown  and  Morris*  state  that  preluninary  washing  with  cold  water  as  in  the 
O'Snllivan  method,  is  unnecessary  in  Tropaeolum  majus. 

**  Sherman  and  Schlesinger,  /.  Am.  Chem.  Soc,  25,  1619  (1913). 


22 

The  injuries  characteristic  of  an  excess  of  each  fertilizer  are  recorded 
from  observations  made  in  the  greenhouse. 

Determinations  of  dry  weight  and  ash  made  upon  the  foHage  of  the 
plants,  showed  an  increase  in  both  values  with  increased  applications  of 
the  fertilizers. 

A  sufficient  number  of  determinations  of  the  mineral  constituents  of 
the  foliage  was  made  to  show  the  increased  content  of  the  fertilizing  salts 
in  the  plants  after  large  applications  of  them  to  the  soil. 

Total  nitrogen  determinations  made  upon  plants  in  different  stages  of 
injury  showed  an  increased  intake  of  nitrogen  when  ammonium  sulfate 
was  applied  but  an  acquired  tolerance  by  the  plant  when  successive  small 
applications  were  made.  Injury  from  ammonium  sulfate  is  not  propor- 
tional to  the  total  nitrogen  content. 

The  sap  was  expressed  from  the  stems  of  the  plants  after  freezing  to 
render  the  plasma  membrane  permeable  to  the  contents  of  the  cells. 
Osmotic  pressure  determinations  made  upon  this  sap  proved  that  with  each 
fertilizer  used  the  degree  of  injury  varied  with  the  osmotic  pressure, 
but  that  not  the  same  degree  of  injury  was  caused  by  different  fertilizers 
at  the  same  osmotic  pressure.  Injury  is  not  a  result  of  increased  osmotic 
pressure  exclusively. 

The  increase  in  the  osmotic  pressure  in  a  series  of  plants  on  soil  receiv- 
ing increasing  applications  of  commercial  fertilizers  was  accompanied  by  an 
increase  in  the  total  solids  and  ash  of  the  sap  and  in  the  amount  of  the 
fertilizer  taken  up  by  the  plant. 

Determinations  of  total  acidity  showed  an  increase  in  the  total  acidity 
of  the  sap  of  plants  fed  with  ammonium  sulfate,  disodium  phosphate  and 
monocalcium  phosphate,  when  phenolphthalein  was  used  as  the  indica- 
tor. 

The  relation  between  the  increase  in  total  acidity  and  in  the  phosphorus 
content  of  the  sap  when  the  plants  were  fed  with  disodium  phosphate 
proved  that  the  phosphorus  was  taken  in  the  form  of  dihydrogen  phos- 
phate, due,  as  was  shown,  not  entirely  at  least  to  absorption  of  the  base 
by  the  soil  but  to  the  selective  action  of  the  plant.  Applications  of  potas- 
sium sulfate  had  no  effect  upon  the  acidity  of  the  sap. 

The  sap  from  the  stems  of  plants  grown  on  soil  to  which  large  applica- 
tions of  potassium  sulfate  had  been  made  showed  a  higher  total  sugar 
content,  the  same  results  being  obtained  with  extracts  of  foliage.  The 
starch  content  of  the  foliage  of  such  plants  was  lower.  These  data  indi- 
cate a  more  rapid  hydrolysis  of  the  starch  in  the  foliage  in  the  presence  of 
an  excess  of  potassium  sulfate.  The  increased  exudation  of  nectar  in  the 
flowers  of  these  plants  probably  resulted  from  this  increase  in  sugar  con- 
tent. 


23 

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2.  Astruc,  A.,  "Rescherches  sur  I'acidite  vegetale,"  Ann.  Sci.  Nat.  Botan.,  [3] 
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6.  Cameron,  F.  K.,  and  Bell,  J.  M.,  "The  Mineral  Constituents  of  the  Soil  Solu- 
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7.  Cameron,  F.  K.,  and  Bell,  J.  M.,  "The  Action  of  Water  and  Aqueous  Solutions 
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8.  Cavara,  F.,  "Resultati  di  una  serie  di  ricerche  crioscopische  sui  vegetali," 
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12.  Drabble,  Eric,  and  Drabble,  Hilda,  "The  Relation  between  the  Osmotic 
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(1907). 

13.  Ewart,  A.  J.,  "The  Ascent  of  Water  in  Trees,"  Trans.  Roy.  Soc.  London,  198, 
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Urbaha,  III. 


",  ^  v5  -^""^  ^* 


BIOGRAPHY. 

The  author  received  his  early  training  in  the  pubhc  schools  of  Paris, 
Illinois  and  in  the  high  schools  of  Paris  and  Flora,  Illinois  and  Terre 
Haute,  Indiana.  He  received  his  Bachelor's  degree  in  Chemistry  from 
Wabash  College  in  1910  and  has  held  the  following  positions  since  that 
time: 

1910-191 1,  Instructor  in  Chemistry  and  Physics,  Urbana,   111.,  High 

School. 
1911-1912,  Assistant  in  Chemistry,  Agricultural  Experiment  Station, 

University  of  Illinois. 
1912-1914,  Assistant  in  Floriculture.  University  of  Illinois. 
1914-1915,  First  Assistant  in  Floricultural  Chemistry,  University  of 
Illinois. 
The  author  is  a  member  of  Sigma  Xi,  Phi  Lambda  Upsilon,  Gamma 
Alpha,  and  the  American  Chemical  Society. 


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